The overall objective of this research proposal will be to determine the role of myocardial Na,K-ATPase inhibition on digitalis action. More specifically this proposal will investigate the following objectives: (1) obtain further evidence that inhibition of the transport enzyme is not responsible for the positive inotropic action of digitalis steroids, (2) attempt to resolve differences in experimental findings between our laboratory and those of other laboratories who show an association of enzyme inhibition with the inotropic action of ouabain, and (3) obtain more definitive evidence to show that inhibition of myocardial Na,K-ATPase may be the mechanism for the cardiotoxic effects of digitalis but not for its positive inotropic action. The following experimental methods will be employed to determine the relationship between Na,K-ATPase inhibition and the inotropic versus cardiotoxic actions of digitalis: (a) correlation of isolated Na,K-ATPase activity with the inotropic action of digitalis following washout in isolated heart experiments, (b) correlation of ouabain inotropy with in vitro dissociation of 3H-ouabain-enzyme complex associated under physiologic ligand binding conditions, and the use of long drug washoff periods to investigate the presence of a second drug dissociation rate constant, and (c) correlation of enzyme inhibition, inotropy, 3H-ouabain dissociation, myocardial Na ion and K ion levels, and electrophysiologic effects of ouabain in arterially-perfused interventricular septal preparations during washout.